Biochemically-significant small molecules (Hofmeister salts, osmolytes, denaturants, collectively referred to as solutes) exert often large, specific effects on fundamental biological processes involving proteins and nucleic acids (e.g. folding, assembly, binding, aggregation). The long term goal of this project is to interpret and predict these effects quantitatively in terms of structural information, and to develop these solutes as probes of interface formation and coupled folding in processes lacking structural information. Insight into the roles of E. coli osmolytes (e.g. Kglutamate, glycine betaine) in the biophysics of regulation of cytoplasmic volume and biopolymer volume fraction in response to osmotic stress will be obtained. Roles of osmolytes in diseases caused by protein misfolding and aggregation in vivo have recently been proposed; these are just one manifestation of what must be widespread, significant effects of changes in type and concentration of intracellular solutes on cellular biochemistry. To achieve these goals, we propose to obtain and interpret quantitative thermodynamic data characterizing interactions of solutes with biopolymer and model surfaces and effects of these solutes on processes. Data will be analyzed using the recently-developed solute partitioning model (SPM), which interprets effects of a solute on a biopolymer process in terms of the change in water accessible surface area of the biopolymer and the local excess or deficit in concentration of the solute in the water of hydration of that surface. The latter is quantified by a microscopic partition coefficient Kp, a property of both the solute and the composition (%nonpolar, polar, charged) of the surface. By determining overall Kp for processes exposing or burying surfaces with widely different compositions, we will build up a database of partition coefficients Kp to predict solute or salt effects on processes where structural data is available. Experiments are proposed to characterize interactions of solutes with uncharged biopolymer surfaces exposed upon unfolding/melting, including circular dichroism studies with marginally-stable proteins (alpha-helical alanine-based peptides and the globular DNA-binding-domain of lac repressor) and UV-melting studies with a marginally stable 12 bp DNA duplex. To interpret these data, osmometric or solubility studies of interactions of these solutes with appropriately chosen model compounds will be performed. In the longer term, Hofmeister salt and solute effects on significant processes burying charged as well as uncharged biopolymer surface (e.g. protein crystallization, protein-nucleic acid interactions) will also be studied. This research will allow effects of Hofmeister salts and solutes, including denaturants GuHCl and urea, osmolytes Kglutamate and glycine betaine, crystallization agents ammonium sulfate, MPD and PEG, and helix stabilizers like TFE, on any protein, nucleic acid or model process to be quantitatively predicted for the first time in terms of structural information. PUBLIC HEALTH RELEVANCE: This research develops a novel and general strategy to measure, interpret and eventually predict effects of small solutes (e.g. denaturants, osmolytes) and noncoulombic effects of Hofmeister salts on protein and nucleic acid processes (e.g. folding, self-assembly, binding, crystallization) in terms of structural information on the amount and composition of the biopolymer surface buried or exposed to water in these processes. By replacing trial and error, the database generated by this research will facilitate the use of solutes and Hofmeister salts to optimize or probe the steps of protein and nucleic acid processes, and will aid in the development of pharmaceutical or bioengineering applications of solutes (e.g. as stabilizers). In addition, these results will help us understand the interactions of intracellular solutes with each other and with biopolymers which are a key determinant of amount of water in the cell, cell volume, biopolymer volume fraction and even growth rate at a given growth condition.